Cervical Cancer Screening via Visual Inspection With Acetic Acid and Lugol Iodine for Triage of HPV-Positive Women

Key Points Question Is it feasible to implement human papillomavirus (HPV) primary screening with visual inspection with acetic acid and Lugol iodine triage in areas with limited health care resources? Findings In this cohort study of 186 863 women in China, an HPV DNA assay was used for primary screening, and visual inspections were used for immediate triage; if both were positive, a referral for colposcopy was conducted within 1 visit. There was a 93.9% completion rate for the triage test and a detection rate of 2.8% for cervical intraepithelial neoplasia grade 2 or higher among HPV-positive individuals. Meaning This study suggests that the implementation of an HPV DNA assay with visual inspection and a colposcopy within 1 visit may mitigate losses to follow-up, detect precancerous lesions, and hold promise for use in low-resource settings.


Introduction
Cervical cancer has posed a heavy disease burden in low-and middle-income countries, where approximately 90% of cases of cervical cancer occur. 1 In China, an estimated 150 700 new cases and 55 700 deaths were attributed to cervical cancer in 2022, 2 with the peak age of cervical cancer incidence occurring between 50 and 54 years. 3The World Health Organization (WHO) proposed screening 70% of women with a high-quality test as a crucial intervention in the pursuit of eliminating cervical cancer. 4Although screening has shown significant success in high-income countries throughout the past few decades, 5,6 the implementation of similar screening programs has faced significant challenges in resource-constrained settings. 7tology has historically been the standard primary screening method and has played a crucial role in reducing cervical cancer mortality.Conversely, in low-resource settings, challenges in promoting cytology stem largely from the limited availability of cytology experts, the high costs, and the need for multistep visits, as required by the cytology-based screening algorithm. 8In 2021, the WHO recommended human papillomavirus (HPV) DNA-based tests as the preferred primary screening approach, 9 which has gained wide acceptance and has robust diagnostic accuracy even in low-resource areas. 10st HPV infections clear spontaneously within a span of 12 to 24 months.As a result, it is essential to perform triage among HPV-positive individuals and refer them to colposcopy for further diagnosis. 11Triage among HPV-positive individuals helps prevent overdiagnosis and unnecessary overtreatment.However, the challenges posed by the high cost, limited availability of adequately trained physicians, and poor adherence to further diagnosis among HPV-positive women in low-resource areas are of concern.As the fifth of 7 algorithms, the WHO recommended visual inspection with acetic acid as an alternative triage method for HPV-positive women to be implemented in low-resource settings where cytology is constrained. 9Visual inspection with acetic acid offers several advantages, including affordability, ease of use, immediate results, and reduced need for recall rounds.Currently, the performance of visual inspection with acetic acid triage among large samples of HPV-positive women has limited supporting evidence. 12,13 2009, a government-led pilot program for cervical cancer screening was initiated in 221 counties in rural China.The program was included in the basic public health services project and expanded to cover the entire female population aged 35 to 64 years in 2019. 14Currently, only 36.8% of women aged 35 to 64 years have ever undergone cervical cancer screening. 15Despite being an upper-and middle-income country, China's challenges in health care access arise due to its large population, uneven development across the central and western regions, and disparities in the distribution of medical resources.As a consequence, accessing medical care becomes difficult for individuals living in areas with limited health resources.This article aims to assess the performance of HPV testing as a primary screening method followed by triage with visual inspection with acetic acid and Lugol iodine for detecting cervical precancerous lesions among women in regions of China with limited health care resources.In addition, the study seeks to use the findings from Ordos City to provide practical insights for other regions facing similar challenges in terms of limited health care resources.
The careHPV test equipment requires only a small area of workspace.It does not require electricity or running water and can be performed rapidly with minimal training required.Therefore, local laboratory technicians in Ordos City conducted careHPV testing at the screening sites in each district or banner.The test is capable of producing accurate results within 2.5 hours.The results of the careHPV testing were HPV negative or HPV positive.
Visual inspection was performed in parallel with the HPV test as cotesting for each woman.
Visual inspection with acetic acid and Lugol iodine involves the examination of the uterine cervix with the naked eye after the application of acetic acid and Lugol iodine, 3% to 5%, which provides simple tests for the early detection of cervical precancerous lesions.The examination does not require any laboratory support, and its result is immediately available.A visual inspection with acetic acid with abnormal results was defined as the observation of thickened white plaques or acetowhite areas usually near the squamocolumnar junction of the cervix.These areas become visible at least 1 minute after the application of acetic acid.A visual inspection with Lugol iodine with abnormal results was defined as bright yellow or mustard yellow areas of the same zone on the cervix, visible after the application of Lugol iodine.If the detected lesion exhibited a cauliflower-like (fungating) mass or ulcer, it was considered suspicious for cancer.The results of the visual inspection were negative, abnormal, or suspected cancer. 9

Colposcopy, Biopsy, and Follow-Up
Visual inspection served as an immediate triage method for HPV-positive women.Women with abnormal visual inspection results were asked to wait half a day for the HPV test results.If women were HPV positive, a colposcopy was conducted during the same visit.On the other hand, women who were diagnosed as HPV positive but who had negative visual inspection results were recalled for rescreening in the subsequent year.Within the framework of this cotesting design, the immediate colposcopy was performed only when women received a diagnosis of suspected cancer on visual inspection, regardless of the HPV test results.
Digital colposcopes (Zonsun Healthcare Co Ltd) were used at each screening site.If the colposcopy results were normal, no biopsy was performed.In cases in which an abnormal colposcopy result occurred and potential lesions were identified, local colposcopists conducted biopsies.The pathology results, which were considered the criterion standard, were classified into different categories: normal or inflammatory, cervical intraepithelial neoplasia 1 (CIN1), CIN2, CIN3, and carcinoma.According to current guidelines in China, women who had CIN2 or higher (CIN2+) confirmed on biopsy were referred to clinical treatment.Women with pathologic findings less than CIN2 were scheduled for a 1-year follow-up.Women who were HPV negative and had normal or abnormal visual inspection results at baseline underwent a 5-year follow-up.

Quality Control and Data Management
Local health care workers were authorized to be involved in our program only if they passed the training before the program started.A database for a regional cervical cancer screening information system was established to gather information from the women who underwent screening, while avoiding duplicate screenings.Data extraction for this study was conducted from the electronic database.The extracted data encompassed women's demographic information, clinical diagnoses from each visit, HPV laboratory test results, and pathology diagnoses.

Statistical Analysis
Statistical analysis was conducted from December 2022 to December 2023.The primary outcomes were the rate of compliance with colposcopy and the detection rate of CIN2+.The parameters listed were calculated and stratified by a 5-year increment age group: (1) number of women screened; (2) number of colposcopies; (3) colposcopy compliance rate; (4) number of histologically confirmed cases of CIN2+ and CIN3+; (5) positive predictive value (PPV) for CIN2+ and CIN3+; (6) number needed to refer to colposcopy to detect 1 case of CIN2+ (or CIN3+): number of women who underwent colposcopies/number of CIN2+ (or CIN3+) identified; and (7) pathologic findings.
The demographic characteristics of the participants were summarized using frequencies and percentages.Differences in outcome distribution among different testing results or age groups were tested using the χ 2 test.All analyses were conducted using SAS, version 9.4 (SAS Institute Inc).All P values were from 2-sided tests and results were deemed statistically significant at P < .05.

Characteristics of the Study Population
Of the 187 863 women (median age, 46 years [IQR, 40-52 years]) who participated in the screening program and had valid HPV results, 24 070 were found to be HPV positive, resulting in an overall HPV positivity rate of 12.8% (Figure).Among the participants, 8557 women refused to undergo visual inspection (1478 HPV positive and 7079 HPV negative).The overall adherence to triage with visual inspection among HPV-positive women was 93.9% (22 592 of 24 070).
Table 1 displays the baseline demographic characteristics of the 179 306 women who had valid results for HPV testing and visual inspection in the study.The median age of this group was 46 years (IQR, 40-52 years).Overall, most women had attained at least a high school degree (60.0% [107 548]), 39 245 (21.9%) were classified as being in menopausal status (median age of menopausal women, 50 years [IQR, 48-53 years]), and most women (60.5% [108 561]) reported not using contraceptives.

Overall Screening Findings
Of 22 592 women who tested positive for HPV with valid visual inspection results, 4094 (18.1%) had abnormal visual inspection results, and 43 (0.2%) had suspected cancer based on visual inspection results.Meanwhile, of 156 714 women who tested negative for HPV, 643 (0.4%) had suspected cancer based on visual inspection.In total, 30.3% of women (1447 of 4780) refused colposcopy.
Among the 4780 women referred to colposcopy, immediate referrals were provided for 686 women with suspected cancer based on visual inspection.The compliance rate for these immediate referrals was high (95.0%[652 of 686]).
Among women who tested positive for HPV, visual inspection reduced colposcopy referral rates to 18.3% (4137 of 22 592) (Table 2).The overall rate of compliance with colposcopy was 65.6% (2714 of 4137).The colposcopy compliance rate was 65.5% (2681 of 4094) among those with abnormal visual inspection results and 76.7% (33 of 43) among those with suspected cancer based on visual inspection.

Principal Findings
To our knowledge, this study is the largest population-based screening to evaluate the performance of HPV testing as the primary screening method with visual inspection as a triage method in the same visit.Our findings showed that among HPV-positive women, the overall adherence to triage by visual inspection was 93.9% (22 592 of 24 070), and the colposcopy compliance rate was 65.6% (2714 of 4137).Among HPV-positive women, visual inspection reduced colposcopy referral rates to 18.3%  Abbreviations: CIN1, cervical intraepithelial neoplasia grade 1; CIN2, cervical intraepithelial neoplasia grade 2; CIN3, cervical intraepithelial neoplasia grade 3. The findings from Ordos City offer valuable insights for areas that face challenges due to limited access to health care resources.

Performance of Visual Inspection Triage of HPV-Positive Women
0][21] According to a meta-analysis, the careHPV test demonstrated a pooled specificity of 84%, 20 implying that it may yield false-positive results for 16 of every 100 women without any abnormalities.Our results suggest that visual inspection may serve as an optional triage test among women whose results of the careHPV test were positive.In comparison with a multicenter, randomized clinical trial conducted rural China in 2015, 22 visual inspection demonstrated a lower positivity rate (18.3% vs 27.1%), which can be attributed to the subjective diagnosis by different physicians. 12,23Meanwhile, a greater detection rate of CIN2+ (2.8% vs 0.5%) and a significant reduction in the number of colposcopy referrals needed to identify 1 case of CIN2+ (4.2 vs 8.0) were observed in our study.These findings highlight the screening efficiency in the practical setting. 22Among women who had abnormal results or suspected cancer based on visual inspection, the PPV for CIN2+ detection was 23.7%, which was slightly lower than the PPV in the ESTAMPA (Estudio Multicéntrico de Tamizaje y Triaje de Cáncer de Cuello Uterino con Pruebas del Virus del Papiloma Humano) study (29.9%). 12This difference could potentially be associated with the inclusion of the results from an 18-month follow-up in the ESTAMPA study, which accumulated more cases of CIN2+.
5][26] Our analysis revealed that triage with visual inspection exhibited a higher detection rate of CIN2+ cases among younger women, which was consistent with previous studies using visual inspection, genotyping, or cytology for HPV-positive triage. 12,27,28This phenomenon could be associated with the retreat of the squamocolumnar junction and the resulting challenges in visualizing endocervical disease in postmenopausal women. 29This inference is further supported by the median age of 50 years of menopausal women in our study.

Concurrent Application of HPV Testing and Visual Inspection
An important advantage associated with triage with visual inspection in our study is the possibility of linking screening to immediate management.Reducing visit numbers is crucial in low-resource areas; for each additional visit, approximately one-fourth to one-third of screened women are lost to follow-up. 30,31Women in these settings often face economic disadvantages and social barriers and encounter limited access to health care services.Failure to provide adequate care to these women may result in a worse prognosis for screening-detectable lesions. 32 Furthermore, the simultaneous application of both tests during the same visit may reduce the possibility of missed diagnoses of CIN2+ among individuals with a negative HPV result because a negative HPV result does not guarantee a risk-free status. 33In our study, we detected a total of 5 cases of cancer among 643 women aged 45 years or older who were HPV negative and received a diagnosis of suspected cancer based on visual inspection.The occurrence of false-negative HPV results can be associated with unsatisfactory sampling among older women, which may result in low sample quality for HPV testing, leading to the inability to detect HPV infection.

Achievement of the Screening Program in Ordos City
Screening coverage is a key metric for monitoring the performance of the WHO cervical cancer elimination plan.The implementation of a sustainable screening program is crucial to achieving coverage, effectively reducing health disparities, and enhancing health equity in low-resource areas. 34Based on our findings, the organized screening program has covered 92.4% of the targeted women and successfully achieved an overall coverage rate of 55.4% among residents within 5 years.
The local government retrospectively collected results conducted at different health care facilities in Ordos City in 2022.After removing duplicates, we identified a total of 76 692 women aged 35 to 64 years who underwent opportunistic screening between 2016 and 2020.Our results suggest that a combination of organized screening and opportunistic screening can achieve a 75.5% screening coverage in a 5-year interval in Ordos City.
To our knowledge, this was the largest study that evaluated the the use of HPV testing for primary screening and visual inspection for triage within the same visit in areas with limited health care resources.The extensive sample size used in this study allows for a comprehensive evaluation.incorporating HPV-16/18 and 12 other genotyping tests. 35Besides screening, Ordos City was the first city in China to implement a regional, free HPV vaccination program for girls aged 13 to 18 years starting in 2021. 36Ordos City is expected to be the first city in China to eliminate cervical cancer.

Limitations
Our study was subject to several limitations.The overall compliance rates for colposcopy recalls were not satisfactory.Approximately 30.3% of women (1447 of 4780) refused colposcopy in local hospitals, which could result in an underestimation of the cumulative detection of CIN2+.This hesitation to undergo colposcopy may be because many women with precancer have no symptoms, and because this is the first-ever free cervical cancer screening by Ordos City government for all women aged 35 to 64 years.The program was implemented in a sparsely populated area, where a subset of women was reluctant to follow the advice of health care professionals to stay and await HPV test results.Nonetheless, if a woman opted for a colposcopy or underwent further diagnosis at a different hospital, the woman was advised to bring back the report of the further examination.

Conclusions
Further investigation is required to explore the cost-effectiveness of the screening approach discussed in this study.Using visual inspection for triage in the same setting in rural China demonstrates higher cost-effectiveness in comparison with cytology for triaging HPV-positive individuals. 37Nonetheless, no studies, to our knowledge, have investigated the economic benefits of triaging HPV-positive individuals using visual inspection while conducting both methods during the same visit.
The findings of this cohort study suggest that integrating HPV testing, visual inspection, and colposcopy in a single visit can effectively reduce loss to follow-up, detect precancerous lesions, and prove to be feasible in large-scale population-based cervical cancer screening programs in

Furthermore, the screening
program underwent regular supervision by experts from the Chinese Academy of Medical Sciences to ensure the quality of the program.Building on the 5 years of experience, Ordos City initiated a new round of HPV-based cervical cancer screening in 2021,

Table 3
presents the age-stratified pathologic findings among women who tested positive for HPV, with abnormal results or suspected cancer based on visual inspection.Table 4 displays the screening findings among age-stratified, HPV-negative women, with suspected cancer results based on visual

Table 1 .
Characteristics of Women With Valid Resultsinspection.A total of 5 cases of cancer were detected.For women who tested positive for HPV and had abnormal results or suspected cancer based on visual inspection, the detection rate of CIN2+ was significantly higher compared with women who were HPV negative with suspected cancer based on visual inspection (15.5% [643 of 4137] vs 2.2% [14 of 643]; χ 2 = 83.8509;P < .001).

Table 2 .
Screening Findings Among Women Who Had careHPV-Positive and Abnormal Results or Suspected Cancer Based on Visual Inspection, Stratified by Age Abbreviations: CIN2+, cervical intraepithelial neoplasia grade 2 or higher; CIN3+, cervical intraepithelial neoplasia grade 3 or higher; NNR, number needed to refer; PPV, positive predictive value.

Table 3 .
Pathologic Findings Among Women With careHPV-Positive and Abnormal Results or Suspected Cancer Based on Visual Inspection, Stratified by Age

Table 4 .
Screening Findings Among Women With careHPV-Negative Results and Suspected Cancer Based on Visual Inspection, Stratified by Age HPV-Positive Triage With Visual Inspection With Acetic Acid and Lugol Iodine (4137 of 22 592).Detection rates were 2.8% (643 of 22 592) for CIN2+ and 1.9% (438 of 22 592) for CIN3+.The results demonstrated that visual inspection with acetic acid and Lugol iodine can stratify risk among HPV-positive individuals, and the implementation of HPV testing, visual inspection, and colposcopy within 1 visit can effectively mitigate losses to follow-up and detect precancerous lesions.